Hydrogen peroxide is used as a potent disinfectant for a broad range of microorganisms, but does not necessarily eliminate all microorganisms during the application time. To check if microorganisms survived the treatment with hydrogen peroxide, suitable growth media are applied. However, remaining hydrogen peroxide inhibits the growth of microorganisms, which leads to false negative results and a false sense of security. Thus hydrogen peroxide needs to be neutralized or degraded in growth media. For this purpose, different additives to growth media have been suggested, such as for example pyruvate or catalase.
Pyruvate reacts with hydrogen peroxide to acetate, carbon dioxide and water. One disadvantage of pyruvate and other chemical additives compared to a catalytic degradation is that the chemical additives are used up by the neutralizing reaction with hydrogen peroxide. Another disadvantage of pyruvate is that microorganisms can metabolize pyruvate. McDonald, et al. (Applied and Environmental Microbiology, 1983, pp 360-365) tested pyruvate addition to violet red bile agar, which is selective for coliforms, and showed that pyruvate lead to false-positive results.
Catalases catalyze the degradation of hydrogen peroxide to the harmless products water and oxygen. Catalases, especially the longest known catalase from bovine liver, have been tried before in growth media, specifically agar plates, with limited success. Calabrese and Bissonnette (Canadian Journal of Microbiology, 1990 pp 544-550) combined bovine liver catalase, 1500 U/plate and sodium pyruvate 5 g/l in agar growth media to increase the recovery of bacteria stressed by acidic water. Ohresser et al. (PDA J Pharm Sci Technol., 2004 pp 75-80) found only 60% recovery with 8000 international units of catalase per plate and stated that “high concentrations of enzyme in the media were not economically viable (in terms of cost per plate)”. Ohresser et al. and McDonald et al. report the unsuccessful application of a catalase in agar plates. These and other reports do not specify the source organism of the catalase, and thus imply that the source of a catalase is not important or it does not make a difference. Harmon and Kautter (Appl Environ Microbiol. 1976 September; 32(3):409-16) describe that catalases from sources other than beef liver were effective in stimulating growth of C. perfringens on different media. The catalase solutions were spread on the surface of the plates because of the low thermal stability of the enzyme.
However, Switala and Loewen (Archives of Biochemistry and Biophysics, 2002 pp 145-154) showed the diversity of properties among catalases, e.g. a broad range of sensitivities to heat inactivation and a wide range of Km values was observed. A catalase with a low value for Km (the calculated Michaelis constant) is desirable in an application with low hydrogen peroxide concentrations.